Selection of aptamers against live bacterial cells

被引:125
作者
Hamula, Camille L. A. [1 ]
Zhang, Hongquan [1 ]
Guan, Le Luo [2 ]
Li, Xing-Fang [1 ]
Le, X. Chris [1 ]
机构
[1] Univ Alberta, Fac Med & Dent, Dept Lab Med & Pathol, Div Analyt & Environm Toxicol, Edmonton, AB T6G 2G3, Canada
[2] Univ Alberta, Dept Agr Food & Nutr Sci, Edmonton, AB T6G 2P5, Canada
基金
加拿大自然科学与工程研究理事会;
关键词
D O I
10.1021/ac801272s
中图分类号
O65 [分析化学];
学科分类号
070302 ; 081704 ;
摘要
Single-stranded DNA or RNA aptamer molecules have usually been selected against purified target molecules. To eliminate the need of purifying target molecules on the cell surface, we have developed a selection technique using live bacterial cells in suspension as targets, to select for ssDNA aptamers specific to cell surface molecules. Lactobacillus acidophilus cells were chosen to demonstrate proof of principle based on their high abundance of surface molecules (potential targets). Aptamer pools obtained after 6-8 rounds of selection demonstrated high affinity for and selective binding with L, acidophilus cells when tested via flow cytometry, microscopy, and fluorescence measurements. Out of 27 aptamers that were cloned and sequenced, one sequence, hemag1P, was found to bind to L. acidophilus much more strongly and specifically than other cells tested. This aptamer was predicted to have a tight hairpin secondary structure. On average, an estimated 164 +/- 47 aptamer molecules were bound to a target cell with an apparent K-d of 13 +/- 3 nM. A likely putative molecular target of hemag1P is the S-layer protein on the cell surface.
引用
收藏
页码:7812 / 7819
页数:8
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